In a flow cytometer, a focused light source such as a laser beam or LED is arranged to shine on particles flowing within a fluid stream. The particles scatter the light and also cause fluorescent light of a different wavelength than the incident beam to be emitted.
The focused spot however is usually not a bright point that decays away monotonically in all directions. Rather, the beam from the light source such as a laser can have “lobes.” The far field of the beam is typically the Fourier transform of the laser aperture. Thus, for example a square aperture will give a far field which is a 2 dimensional Sync function as shown in FIG. 1. The beam shaping optical elements may also have small reflections from various surfaces as shown in FIG. 3, coatings can reduce the reflections but typically will not completely eliminate them. Both of these effects have the net effect of causing the laser beam in the flow channel 17 to have a main peak 3 flanked on one or both sides by many smaller peaks 4 as shown in FIG. 3 and FIG. 2.
While the power in a laser minor side lobe 4 is typically 1% or less of the main lobe 3, a difficulty arises when very large particles 20 and very small particles 21 as shown in FIG. 2 are mixed. A large particle passing over a minor lobe of the laser will cause an amount of signal that is comparable to a small particle that is passing the main lobe of the laser. A large particle passing over a minor lobe of the laser and then subsequently passing over the main lobe of the laser can incorrectly be counted as a small particle followed by a large particle.